Identification of microRNAs in PCV2 subclinically infected pigs by high throughput sequencing

Abstract

Porcine circovirus type 2 (PCV2) is the essential etiological infectious agent of PCV2-systemic disease and has been associated with other swine diseases, all of them collectively known as porcine circovirus diseases. MicroRNAs (miRNAs) are a new class of small non-coding RNAs that regulate gene expression post-transcriptionally. miRNAs play an increasing role in many biological processes. The study of miRNA-mediated host-pathogen interactions has emerged in the last decade due to the important role that miRNAs play in antiviral defense. The objective of this study was to identify the miRNA expression pattern in PCV2 subclinically infected and non-infected pigs. For this purpose an experimental PCV2 infection was carried out and small-RNA libraries were constructed from tonsil and mediastinal lymph node (MLN) of infected and non-infected pigs. High throughput sequencing determined differences in miRNA expression in MLN between infected and non-infected while, in tonsil, a very conserved pattern was observed. In MLN, miRNA 126-3p, miRNA 126-5p, let-7d-3p, mir-129a and mir-let-7b-3p were up-regulated whereas mir-193a-5p, mir-574-5p and mir-34a down-regulated. Prediction of functional analysis showed that these miRNAs can be involved in pathways related to immune system and in processes related to the pathogenesis of PCV2, although functional assays are needed to support these predictions. This is the first study on miRNA gene expression in pigs infected with PCV2 using a high throughput sequencing approach in which several host miRNAs were differentially expressed in response to PCV2 infection.Electronic supplementary materialThe online version of this article (doi:10.1186/s13567-014-0141-4) contains supplementary material, which is available to authorized users.

Evaluation of the capability of the PCV2 genome to encode miRNAs: lack of viral miRNA expression in an experimental infection

“…We have reported previously modification of the expression pattern of host miRNAs due to the PCV2 infection [ 18 ]. For both purposes, four animals were inoculated with 7 × 10 4.8 TCID 50 of PCV2 isolate Sp-10-7-54-13 [ 19 ] and two animals were inoculated with PBS.…”

“…The analysis of the reads that aligned to the Sus scrofa genome are reported elsewhere because these sequences constitute porcine miRNAs [ 18 ]. For viral miRNA discovery, sequences were blasted to the PCV2 isolate Sp-10-7-54-13 genome (NCBI Reference Sequence: GU049342) considering only sequences with 100% of alignment and identity (perfect match).…”

“…In a previous study [ 18 ], in subclinically infected pigs, we demostrated that PCV2 can alter the miRNA expression pattern of the host. If PCV2 can express miRNAs in vitro has to be determined, but in its natural host, in a subclinical infection, NGS failed to indentify viral miRNAs.…”

Interactions of porcine circovirus 2 with its hosts

“…A further study elucidated that both miR-139-5p and let-7e are down-regulated by the Rep protein [78]. During PCV2 infection, cellular miRNAs in mediastinal lymph nodes were involved in the immune system and in PCV2 pathogenesis [79]. In contrast, viral miRNAs were also reported to participate in viral or host cellular gene expression [80].…”

“…Cellular microRNA (miRNA) pathways play key roles in controlling virus-host interactions [78,79]. Hong and colleagues identified 157 PCV2-regulated miRNAs in PCV2-infected PK15 cells, most of which function as posttranscriptional regulators and MAPK signal activators during PCV2 infection [78].…”